Post-partum mammalian placenta, its use and placental stem cells therefrom

• US 7,255,879 B2
• Filed: 08/12/2003
• Issued: 08/14/2007
• Est. Priority Date: 12/06/2000
• Status: Expired due to Term

• Current Assignee: Celularity, Inc.
• Original Assignee: Anthrogenesis Corporation (Bristol-Myers Squibb Company)
• Inventors: Hariri, Robert J.
• Primary Examiner(s): Li; Q. Janice
• Application Number:
  US10/640,428
• Publication Number:
  US 20040048372A1
• Time in Patent Office: 1,463 Days
• Field of Search: None
• US Class Current: 424/529
• CPC Class Codes: A61K 35/12 Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells vaccines or medicinal preparations containing antigens or antibodies A61K39/00
• CPC Class Codes: A61K 48/00 Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
• CPC Class Codes: A61M 1/02 Blood transfusion apparatus blood infusion by syringes A61M5/14
• CPC Class Codes: A61M 2202/0462 Placental blood, umbilical cord blood
• CPC Class Codes: A61M 2210/1458 Placenta
• CPC Class Codes: A61M 2210/1466 Umbilical cord
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/16 Anti-Parkinson drugs
• CPC Class Codes: A61P 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• CPC Class Codes: A61P 27/06 Antiglaucoma agents or miotics
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 5/14 of the thyroid hormones, e.g. T3, T4
• CPC Class Codes: A61P 7/06 Antianaemics
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/10 : for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: C12N 2500/90 : Serum-free medium, which may still contain naturally-sourced components
• CPC Class Codes: C12N 2509/00 : Methods for the dissociation of cells, e.g. specific use of enzymes
• CPC Class Codes: C12N 2533/90 : Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
• CPC Class Codes: C12N 5/0068 : General culture methods using substrates for specific animal cell type C12N5/06
• CPC Class Codes: C12N 5/0605 : Cells from extra-embryonic tissues, e.g. placenta, amnion, yolk sac, Wharton's jelly

Abstract

The present invention provides a method of extracting and recovering embryonic-like stem cells, including, but not limited to pluripotent or multipotent stem cells, from an exsanguinated human placenta. A placenta is treated to remove residual umbilical cord blood by perfusing an exsanguinated placenta, preferably with an anticoagulant solution, to flush out residual cells. The residual cells and perfusion liquid from the exsanguinated placenta are collected, and the embryonic-like stem cells are separated from the residual cells and perfusion liquid. The invention also provides a method of utilizing the isolated and perfused placenta as a bioreactor in which to propagate endogenous cells, including, but not limited to, embryonic-like stem cells. The invention also provides methods for propagation of exogenous cells in a placental bioreactor and collecting the propagated exogenous cells and bioactive molecules therefrom.

Claims

1. A method of collecting a population of cells comprising human CD34⁻

placental stem cells from post-partum exsanguinated human placenta, said method comprising;

perfusing said placenta with a perfusion solution in an amount and for a time sufficient to collect cells, wherein said placenta has been drained of cord blood and flushed to remove residual blood prior to said perfusing, and wherein said perfusing is performed by passing said perfusion solution into one or both of the umbilical artery and umbilical vein of said placenta; and

collecting said perfusion solution from said placenta so that said population of cells is collected, wherein said population of cells comprises CD34⁻

placental stem cells.

2. The method of claim 1, wherein said perfusing is performed at least four hours after removal of said residual blood.

3. The method of claim 1, wherein said perfusing is performed at least twelve hours after removal of said residual blood.

4. The method of claim 1, wherein said perfusing is performed at least 24 hours after removal of said residual blood.

5. The method of claim 1, wherein said perfusing is performed using a first volume of between about 30 ml and about 150 ml of said perfusion solution.

6. The method of claim 5, further comprising continuing said perfusing using a second volume of about 30 ml to about 150 ml of said perfusion solution, said second volume being collected separately from said first volume.

7. The method of claim 1, in which said perfusing is performed for a plurality of times.

8. The method of claim 7, wherein, for each of said times, said perfusing is performed using a volume of about 30 ml to about 150 ml of said perfusion solution.

9. The method of claim 1, wherein said perfusion solution contains an anticoagulant.

10. The method of claim 5, wherein said perfusion solution contains an anticoagulant.

11. The method of claim 7, wherein said perfusion solution contains an anticoagulant.

12. The method of claim 1, wherein said perfusion solution comprises heparin, ethylene diamine tetra acetic acid (EDTA) or creatine phosphate dextrose (CPDA).

13. The method of claim 1, wherein said perfusion solution comprises a growth factor or a cytokine.

14. The method of claim 13, wherein said growth factor or cytokine is selected from the group consisting of a colony stimulating factor, interferon, erythropoietin, stem cell factor, thrombopoietin, an interleukin, granulocyte colony-stimulating factor, and any combination thereof.

15. The method of claim 1, wherein said human placenta is a post-partum placenta remaining after a successful birth.

16. The method of claim 1, wherein said CD34⁻

stem cells are multipotent.

17. The method of claim 1, wherein said population of cells are collected from a placenta that has been maintained for up to 2 hours at room temperature.

18. The method of claim 1, wherein said population of cells are collected from a placenta that has been maintained for up to 12 hours at room temperature.

19. The method of claim 1, wherein said population of cells are collected from a placenta that has been maintained for up to 48 hours at room temperature.

20. The method of claim 7, wherein said population of cells are collected from a placenta that has been maintained for up to 2 hours at room temperature.

21. The method of claim 7, wherein said population of cells are collected from a placenta that has been maintained for up to 12 hours at room temperature.

22. The method of claim 7, wherein said population of cells are collected from a placenta that has been maintained for up to 48 hours at room temperature.

23. The method of claim 1, wherein said CD34⁻

stem cells are positive for SH2, SH3 or SH4.

24. The method of claim 1, wherein said CD34⁻

stem cells are negative for CD45, SSEA3 or SSEA4.

25. The method of claim 1, wherein said CD34⁻

stem cells are positive for CD10, CD29, CD44, CD54, CD90, or OCT-4.

26. The method of claim 1, wherein said CD34⁻

stem cells are CD10⁺, CD29⁺, CD44⁺, CD45⁻

, CD54⁺, CD90⁺, SH2⁺, SH3⁺, SH4⁺, SSEA3⁻

, SSEA4⁻

, and OCT-4⁺.